Abstract

The abundance of the multimeric vacuolar ATP dependent proton pump, V-ATPase, on the plasma membrane of tumor cells correlates with the invasiveness of the tumor cell, suggesting the involvement of V-ATPase in tumor metastasis. V-ATPase is hypothesized to create a proton efflux leading to an acidic pericellular microenvironment that promotes the activity of pro-invasive proteases. An alternative, not yet explored possibility is that V-ATPase regulates the signaling machinery responsible for tumor cell migration. Here, we show that pharmacological or genetic reduction of V-ATPase activity significantly reduces migration of invasive tumor cells in vitro. Importantly, the V-ATPase inhibitor archazolid, abrogates tumor dissemination in a syngeneic mouse 4T1 breast tumor metastasis model. Pretreatment of cancer cells with archazolid impairs directional motility by preventing spatially-restricted, leading edge localization of epidermal growth factor receptor (EGFR) as
well as of phosphorylated Akt. Archazolid treatment or silencing of V-ATPase inhibited Rac1 activation, as well as Rac1-dependent dorsal and peripheral ruffles by inhibiting Rab5-mediated endocytotic/exocytotic trafficking of Rac1. The results indicate that archazolid effectively decreases metastatic dissemination of breast tumors by impairing the trafficking and spatially-restricted activation of EGFR and Rho-GTPase Rac1, which are pivotal for directed movement of cells. Thus, our data reveals a novel mechanism underlying the role of V-ATPase in tumor dissemination.